QUANTITATIVE TRANSIENT GENE-EXPRESSION - COMPARISON OF THE PROMOTERS FOR MAIZE POLYUBIQUITIN1, RICE ACTIN1, MAIZE-DERIVED EMU AND CAMV 35S IN CELLS OF BARLEY, MAIZE AND TOBACCO

The particle gun approach was used for the quantification of promoter efficiency in a test system for transient gene expression. Beta-glucur onidase was used as reporter gene for determining promoter strength. T he variability inherent in this gene transfer system was considerably reduced by calculating a transformation efficiency factor given by the expression of a cotransferred second reporter gene (firefly luciferas e). The calibration of beta-glucuronidase activity by the transformati on efficiency factor caused a lower statistical variance of the values and allowed reliable results to be obtained with a smaller set of rep etitions. The CaMV 35S promoter (as a control) and the monocot-specifi c promoters for maize polyubiquitin1, rice actin1 and the maize-derive d Emu were characterized and compared with respect to expression stren gth, as tested under identical conditions in suspension cell cultures of maize, barley and tobacco. Compared to the 35S promoter, the monoco t-specific promoters show up to 15-fold higher expression in maize and barley but give only weak expression in tobacco. No expression was fo und for the rice actin1 promoter in tobacco. The level of reporter gen e expression is influenced by the osmotic potential in the agar medium . For the Emu promoter, the calibrated beta-glucuronidase activities r emained nearly constant at low sucrose concentrations. Above 8% sucros e, the calibrated activities increased steadily with increasing osmoti c conditions, reaching a three- to four-fold higher level at the highe st sucrose concentration (32%) as compared to the standard concentrati on (4% sucrose) in the medium.